Project Summary. Our objective is to develop small molecule inhibitors of the highly oncogenic protein tyrosine phosphatase, PTP4A3, as a targeted therapy for ovarian cancer (OvCa). Elevated levels of PTP4A3 mRNA and protein in ovarian tumors correlate with disease progression and recurrence, poor patient prognosis and poor patient survival. Genetic depletion of PTP4A3 in cancer cells diminishes their ability to survive, migrate and form tumors in vivo. Conversely, PTP4A3 overexpression increases tumor cell migration, invasion and dissemination in multiple cancers, including OvCa. Taken together, these data suggest PTP4A3 is a novel oncogenic molecular target for OvCa. Currently, there are no small molecule PTP4A3 inhibitors in clinical development for any type of cancer. We discovered the most potent known in vitro small molecule inhibitor of PTP4A3, which has a Ki of 18 nM (JMS-053; 7-imino-2- phenylthieno[3,2-c]pyridine-4,6(5H,7H)-dione). JMS-053 displayed no significant in vitro inhibition of 21 other protein phosphatases and 49 kinases at 1 M, suggesting considerable specificity towards PTP4A3. JMS-053 killed OvCa cells grown as 3D spheroids, including those derived from high grade serous and drug resistant OvCa cell lines, with EC50 values as low as 600 nM. JMS-053 treatment also block the migration of OvCa cells and decreased RhoA activity. JMS-053 did not inhibit the growth of the human ovarian surface epithelial cells at concentrations at least up to 25 M. JMS-053 (10 mg/kg) displayed anticancer activity in a murine xenograft model of drug resistant OvCa. To improve the pharmaceutical properties of JMS-053, we have synthesized next generation analogs that have superior IC50 values for PTP4A3 in vitro and are computationally more drug-like. KeViRx proposes to credential these analogs to identify which compound(s) should be developed further as potential targeted therapeutics for OvCa. We propose three Tasks: Task 1. Define the single agent actions of JMS-053 analogs in drug-sensitive and -resistant human OvCa in vitro. Task 2. Determine the interactions of JMS-053 and its analogs with clinically approved drugs for the treatment of OvCa in vitro. Task 3. Investigate the maximum tolerated in vivo dose, the pharmacokinetics and preliminary antitumor efficacy of two selected JMS-053 analogs. These studies should position at least one small molecule PTP4A3 inhibitor for further Phase II SBIR funded development in GMP-grade analog studies.